Pre-existing polymerase-specific T cells expand in abortive seronegative SARS-CoV-2
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (ScreenIT)
Abstract
Individuals with potential exposure to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) do not necessarily develop PCR or antibody positivity, suggesting that some individuals may clear subclinical infection before seroconversion. T cells can contribute to the rapid clearance of SARS-CoV-2 and other coronavirus infections 1–3 . Here we hypothesize that pre-existing memory T cell responses, with cross-protective potential against SARS-CoV-2 (refs. 4–11 ), would expand in vivo to support rapid viral control, aborting infection. We measured SARS-CoV-2-reactive T cells, including those against the early transcribed replication–transcription complex (RTC) 12,13 , in intensively monitored healthcare workers (HCWs) who tested repeatedly negative according to PCR, antibody binding and neutralization assays (seronegative HCWs (SN-HCWs)). SN-HCWs had stronger, more multispecific memory T cells compared with a cohort of unexposed individuals from before the pandemic (prepandemic cohort), and these cells were more frequently directed against the RTC than the structural-protein-dominated responses observed after detectable infection (matched concurrent cohort). SN-HCWs with the strongest RTC-specific T cells had an increase in IFI27 , a robust early innate signature of SARS-CoV-2 (ref. 14 ), suggesting abortive infection. RNA polymerase within RTC was the largest region of high sequence conservation across human seasonal coronaviruses (HCoV) and SARS-CoV-2 clades. RNA polymerase was preferentially targeted (among the regions tested) by T cells from prepandemic cohorts and SN-HCWs. RTC-epitope-specific T cells that cross-recognized HCoV variants were identified in SN-HCWs. Enriched pre-existing RNA-polymerase-specific T cells expanded in vivo to preferentially accumulate in the memory response after putative abortive compared to overt SARS-CoV-2 infection. Our data highlight RTC-specific T cells as targets for vaccines against endemic and emerging Coronaviridae .
Article activity feed
-
-
SciScore for 10.1101/2021.06.26.21259239: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IRB: COVIDsortium Healthcare Worker Participants: The COVIDsortium bioresource was approved by the ethical committee of UK National Research Ethics Service (20/SC/0149) and registered on ClinicalTrials.gov (NCT04318314).
Consent: All subjects gave written informed consent and the study conformed to the principles of the Helsinki Declaration.Sex as a biological variable not detected. Randomization Other experiments were not randomized and the investigators were not blinded to allocation during experiments and outcome assessment. Blinding Other experiments were not randomized and the investigators were not blinded to allocation during experiments and outcome assessment. Power Analysis Power … SciScore for 10.1101/2021.06.26.21259239: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IRB: COVIDsortium Healthcare Worker Participants: The COVIDsortium bioresource was approved by the ethical committee of UK National Research Ethics Service (20/SC/0149) and registered on ClinicalTrials.gov (NCT04318314).
Consent: All subjects gave written informed consent and the study conformed to the principles of the Helsinki Declaration.Sex as a biological variable not detected. Randomization Other experiments were not randomized and the investigators were not blinded to allocation during experiments and outcome assessment. Blinding Other experiments were not randomized and the investigators were not blinded to allocation during experiments and outcome assessment. Power Analysis Power calculations were performed prior to week 16 sub-study sampling to determine the sample size needed to test the hypothesis that HCW with pre-existing T cell responses are enriched in exposed uninfected group at a range of incidence of infection, assuming 50% of cohort had pre-existing T cell responses. Cell Line Authentication Authentication: Spike ELISA: Seropositivity against SARS-CoV-2 spike was determined for medical student and laboratory staff cohort between July 2020 and Jan 2021 (Extended Data Table 4) by enzyme-linked immunosorbent assay, as validated and described previously50,60,61. Table 2: Resources
Antibodies Sentences Resources Lab confirmed infection was determined by weekly nasopharyngeal RNA stabilizing swabs and reverse transcriptase polymerase chain reaction (RT-PCR; Roche cobas SARS-CoV-2 test, Envelope [E] gene) and antibody assay positivity (Spike protein 1 IgG Ab assay, EUROIMMUN) and anti-nucleocapsid total antibody assay (ROCHE) described in detail below. Spike protein 1 IgGsuggested: Noneanti-nucleocapsid totalsuggested: NoneELISpot plates were developed with human biotinylated IFN-γ detection antibody (7-B6-1, Mabtech; 1μg/ml) for hr at RT, followed by incubation with goat anti-biotin alkaline phosphatase (Vector Laboratories; 1:1000) for 2 hr RT, both diluted in PBS with 0.5% BSA by volume (Sigma-Aldrich), and finally with 50 μl/well of sterile filtered BCIP/NBT Phosphatase Substrate (ThermoFisher) for 7 min RT. anti-biotin alkaline phosphatasesuggested: NoneCells were washed in PBS and incubated in perm buffer (TF staining buffer kit, diluted 1:10 in ddH2O) for 20 mins RT, washed in PBS and resuspended in perm buffer with saturating concentrations of anti-human antibodies for intracellular staining: IL-2 PerCp-eFluor710 (Invitrogen, clone MQ1-17H12) anti-humansuggested: NoneCTV dilution and staining with anti-human-IFNγ antibodies was used to identify antigen-specific T cells. anti-human-IFNγsuggested: NoneExperimental Models: Cell Lines Sentences Resources 4 × 104 Huh7 cells suspended in 100 μl complete media were added per well and incubated for 72 hr at 37 °C and 5% CO2. Huh7suggested: NoneAfter initial incubation, pre-seeded Vero E6 cells were infected with the serum-virus samples and incubated (37 °C and 5% CO2) for 72 hr. Vero E6suggested: NoneSoftware and Algorithms Sentences Resources COVIDsortium Healthcare Worker Participants: The COVIDsortium bioresource was approved by the ethical committee of UK National Research Ethics Service (20/SC/0149) and registered on ClinicalTrials.gov (NCT04318314). COVIDsortium Healthcaresuggested: NoneThe curves of relative infection rates (in %) versus the serum dilutions (log10 values) against a negative control of pooled sera collected prior to 2016 (Sigma) and a positive neutraliser were plotted using Prism 9 (GraphPad). GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)ODs were measured using a MultiskanFC (Thermofisher Scientific) plate reader at 405 nm and S1-specific IgG titers interpolated from the IgG standard curve using 4PL regression curve-fitting on GraphPad Prism 8. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)Brilliant Violet Buffer (BD Biosciences): CD3 Bv510 (Biolegend, clone OKT3) BD Biosciences)suggested: NoneData were analysed by FlowJo version 10.7 (TreeStar) FlowJosuggested: (FlowJo, RRID:SCR_008520)A protein BLAST search of each 15mer peptide was then performed against a custom sequence database comprising 2531 Coronaviridae sequences40. BLASTsuggested: (BLASTX, RRID:SCR_001653)Sequences were aligned using the MUSCLE algorithm with default parameters and percentage identity was calculated in Geneious Prime 2020.1.2 ( MUSCLEsuggested: (MUSCLE, RRID:SCR_011812)(https://www.geneious.com). https://www.geneious.comsuggested: (Geneious, RRID:SCR_010519)Alignment figures were made in Snapgene 5.1 (GSL Biotech). qPCR: Total RNA from Tempus blood was extracted using the Tempus Spin RNA isolation kit (Applied Biosystems, 4380204). cDNA was obtained using the High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems). Snapgenesuggested: (SnapGene, RRID:SCR_015052)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: We found the following clinical trial numbers in your paper:
Identifier Status Title NCT04318314 Recruiting COVID-19: Healthcare Worker Bioresource: Immune Protection a… Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
Results from scite Reference Check: We found no unreliable references.
-
